Multiplex PCR for detection of bacterial uropathogens in nosocomial samples from northeastern Mexico: One Health relevant urinary tract pathogens
DOI:
https://doi.org/10.66585//ohmi.2026.2.0025Keywords:
Mexico-USA border region, Multiplex PCR, Nosocomial surveillance, One Health approach, UropathogensAbstract
Urinary tract infections (UTIs) are among the most common bacterial infections worldwide, accounting for approximately 150 million cases annually, including an estimated 3.8 million in Mexico. The principal uropathogens are Escherichia coli, Klebsiella pneumoniae, and Proteus mirabilis, all of which are opportunistic zoonotic bacteria associated with humans, companion animals, livestock, and wildlife. Within the One Health framework, these pathogens and their antimicrobial resistance (AMR) determinants may circulate across healthcare settings, communities, animals, and the environment, highlighting the need for integrated surveillance. Although conventional urine culture remains the diagnostic gold standard, rapid molecular methods can facilitate timely diagnosis and support antimicrobial stewardship. This study analyzed 300 urine samples collected from three hospitals representing different levels of healthcare complexity in Reynosa, Tamaulipas, Mexico. Conventional urine culture and species-specific PCR assays targeting E. coli, K. pneumoniae, and P. mirabilis were performed. Fifty-six samples yielded positive urine cultures, with E. coli identified as the predominant uropathogen (85.7%, 48/56), followed by K. pneumoniae (10.7%, 6/56) and P. mirabilis (3.6%, 2/56). Positive cases were more frequent among females (70%). In addition, a multiplex PCR assay was developed and validated to simultaneously detect the three pathogens. The assay was also successfully applied directly to unprocessed urine samples, provided that urine represented no more than 20% of the total PCR reaction volume. The multiplex PCR demonstrated 100% concordance with conventional culture results while substantially reducing diagnostic turnaround time. To our knowledge, this is the first study conducted in Reynosa, Tamaulipas, to integrate conventional microbiological and molecular diagnostics for UTIs caused by One Health bacterial pathogens. These findings establish a methodological baseline for regional surveillance of uropathogens at the human–healthcare–environment interface, supporting future cross-border One Health surveillance initiatives.
References
1. Sheerin NS. Urinary tract infection. Medicine. 2015;43(8):435-439. https://doi.org/10.1016/j.mpmed.2015.05.007
2. Nicolle LE, Gupta K, Bradley SF, Colgan R, DeMuri GP, Drekonja D, et al. Clinical practice guideline for the management of asymptomatic bacteriuria: 2019 update by the Infectious Diseases Society of America. Clin Infect Dis. 2019;68(10):83-110. https://doi.org/10.1093/cid/ciy1121
3. Palusiak A. Proteus mirabilis and Klebsiella pneumoniae as pathogens capable of causing co-infections and exhibiting similarities in their virulence factors. Front Cell Infect Microbiol. 2022;12:991657. https://doi.org/10.3389/fcimb.2022.991657
4. Price LB, Hungate BA, Koch BJ, Davis GS, Liu CM. Colonizing opportunistic pathogens (COPs): The beasts in all of us. PLoS Pathog. 2017;13(8):1006369. https://doi.org/10.1371/journal.ppa
t.1006369
5. Jakobsen L, Garneau P, Bruant G, Harel J, Olsen SS, Porsbo LJ, et al. Is Escherichia coli urinary tract infection a zoonosis? Proof of direct link with production animals and meat. Eur J Clin Microbiol Infect Dis. 2012;31(6):1121-1129. https://doi.org/10.1007/s10096-011-1417-5
6. Benoit SR, Ellingson KD, Waterman SH, Pearson ML. Antimicrobial resistance in eight US hospitals along the US-Mexico border, 2000-2006. Epidemiol Infect. 2014;142(11):2378-2387. https://doi.org/1
0.1017/S095026881300318X
7. Pezzoli K, Kozo J, Ferran K, Wooten W, Rangel Gomez G, Al-Delaimy WK. One Bioregion/One Health: An integrative narrative for transboundary planning along the US-Mexico border. Glob Soc. 2014;28(4):419-440. https://doi.org/10.1080/13600826.2014.951316
8. Kumar MS, Ghosh S, Nayak S, Das AP. Recent advances in biosensor-based diagnosis of urinary tract infection. Biosens Bioelectron. 2016;80:497-510. https://doi.org/10.1016/j.bios.2016.02.023
9. Schrick L, Nitsche A. Pitfalls in PCR troubleshooting: Expect the unexpected? Biomol Detect Quantif. 2016;6:1-3. https://doi.org/10.
1016/j.bdq.2015.11.001
10. Villalobo E, Torres A. PCR for detection of Shigella spp. in mayonnaise. Appl Environ Microbiol. 1998;64(4):1242-1245. https://doi.org/10.1128/AEM.64.4.1242-1245.1998
11. Clinical and Laboratory Standards Institute. Performance standards for antimicrobial susceptibility testing. 34th ed. CLSI supplement M100. Wayne (PA). Clinical and Laboratory Standards Institute. 2024.
12. Green MR, Sambrook J. Molecular cloning: A laboratory manual. 4th ed. Cold Spring Harbor (NY). Cold Spring Harbor Laboratory Press. 2012.
13. Flores-Mireles AL, Walker JN, Caparon M, Hultgren SJ. Urinary tract infections: Epidemiology, mechanisms of infection and treatment options. Nat Rev Microbiol. 2015;13(5):269-284. https://doi.org/10.1038/nrmicro3432
14. Hooton TM. Clinical practice. Uncomplicated urinary tract infection. N Engl J Med. 2012;366(11):1028-1037. https://doi.org/10.1056/
NEJMcp1104429
15. Bonkat G, Kranz J, Cai T, Geerlings SE, Köves B, Lambregts MMC, et al. EAU guidelines on urological infections. Arnhem (The Netherlands). EAU Guidelines Office. 2026. https://uroweb.org/gui
delines/urological-infections
16. Schmitz JE, Stratton CW, Persing DH, Tang YW. Forty years of molecular diagnostics for infectious diseases. J Clin Microbiol. 2022;60(10):02446-21. https://doi.org/10.1128/jcm.02446-21
17. Baimakhanova B, Sadanov A, Berezin V, Baimakhanova G, Trenozhnikova L, Orasymbet S, et al. Emerging technologies for the diagnosis of urinary tract infections: Advances in molecular detection and resistance profiling. Diagnostics. 2025;15(19):2469. https://doi.org/10.3390/diagnostics15192469
18. Hao X, Cognetti M, Patel C, Jean-Charles N, Tumati A, Burch-Smith R, et al. The essential role of PCR and PCR panel size in comparison with urine culture in identification of polymicrobial and fastidious organisms in patients with complicated urinary tract infections. Int JMolSci.2023;24(18):14269. https://doi.org/10.3390/ijms241814269
19. Wojno KJ, Baunoch D, Luke N, Opel M, Korman H, Kelly C, et al. Multiplex PCR based urinary tract infection (UTI) analysis compared to traditional urine culture in identifying significant pathogens in symptomatic patients. Urology. 2020;136:119-126. https://doi.org/10.1016/j.urology.2019.10.018
20. Huggett JF, Novak T, Garson JA, Green C, Morris-Jones SD, Miller RF, et al. Differential susceptibility of PCR reactions to inhibitors: an important and unrecognised phenomenon. BMC Res Notes. 2008;1:70. https://doi.org/10.1186/1756-0500-1-70
21. Nicholson EB, Concaugh EA, Foxall PA, Island MD, Mobley HLT. Proteus mirabilis urease: Transcriptional regulation by UreR. J Bacteriol. 1993;175(2):465-473. https://doi.org/10.1128/jb.175.2.
465-473.1993
22. Armbruster CE, Mobley HLT. Merging mythology and morphology: The multifaceted lifestyle of Proteus mirabilis. Nat Rev Microbiol. 2012;10(11):743-754. https://doi.org/10.1038/nrmicro2890
23. Eissa N, Salman MB, Younes AM, Mohamed ESA, Abu-Seida AM, Abdulkarim A, et al. One Health approach on zoonotic multidrug-resistant Klebsiella pneumoniae isolated from Egyptian cattle, horses, and humans. Open Vet J. 2025;15(9):4219-4234. https://doi.org/10.5455/OVJ.2025.v15.i9.28
24. Hasona IF, Awad A, Younis G, Mohamed WF. A One Health perspective on Proteus mirabilis: The interaction of virulence and antimicrobial resistance across human and animal reservoirs. Microorganisms. 2026;14(2):444. https://doi.org/10.3390/microorganisms14020444
25. Ramatla T, Mafokwane T, Lekota K, Monyama M, Khasapane G, Serage N, et al. “One Health” perspective on prevalence of co-existing extended-spectrum β-lactamase (ESBL)-producing Escherichia coli and Klebsiella pneumoniae: A comprehensive systematic review and meta-analysis. Ann Clin Microbiol Antimicrob. 2023;22(1):88. https://doi.org/10.1186/s12941-023-00638-3
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